Pathological exon skipping in an HNPCC proband with MLHI splice acceptor site mutation

One of the most commonly mutated mismatch repair genes in human nonpolyposi s colorectal cancer (HNPCC) is MLHI. We identified a splice site mutation i n MLHI in a colorectal cancer proband (T-to-A at position -11 of intron I s plice acceptor) and investigated its functional consequences by RT-PCR, usi ng lymphocyte mRNA from the proband, two noncarrying siblings, and one unre lated individual. Subcloning of PCR products followed by sequencing of indi vidual clones revealed increased transcript heterogeneity in the mutation c arrier, attributable to the presence of a variety of mRNA forms lacking exo n 2, or combinations of exons 2, 4, 6, 9, and 10. The full-length transcrip t subcloned from the mutation carrier was detected with a much reduced freq uency, suggesting that only the wild-type allele produced functional MLHI m RNA. The three noncarriers expressed some previously described transcripts and several novel variants, but none that lacked exon 2. The results are co nsistent with the hypothesis that this splice site mutation causes skipping of MLHI exon 2 in a large proportion of mRNA transcripts derived from the mutated allele. Such an observation strengthens the case for identifying th e mutation as pathogenic in this HNPCC family, which is of interest given t he rarity of exon skipping defects resulting from splice acceptor site muta tions outside the invariant AG dinucleotide. (C) 2000 Wiley-Liss, Inc.